Allograft rejection is mediated by the recipient's immunological response to donor antigen, initiated and coordinated by CD4+ T cells. Once T cells encounter alloantigen, they undergo expansion and differentiation into effectors and/or memory T cells. Activation also results in the expansion of regulatory T cells that function to control the immune response, and it is proposed that this process of immunoregulation is critical for long term allograft survival. Within T cells, several adapter proteins have been found to play central roles in T cell receptor-induced signal transduction, and several have been found to be active participants in the formation of signaling complexes, which modulate the T cell activation response. T-cell-Specific Adaptor Protein (TSAd) is a SH2 domain containing intracellular adaptor molecule that was initially reported to be restricted in its expression to T cells. Increasing evidence suggests that the function of TSAd in T cells is complex, and that it is of importance in both effector as well as well as regulatory responses. However, no study has addressed questions about the biology of TSAd in allograft rejection and/or in the alloimmune response. This exploratory research proposal is based on novel preliminary observations in which we observed that TSAd knockout recipients of cardiac allografts have a profound defect in alloimmune regulation. The overall goal of our research is to characterize how TSAd mediates immunoregulation following transplantation. Specifically, we plan to use TSAd knockout mice as recipients of allografts to determine its function in the rejection process, to identify its role in the generation and function of alloreactive T cells and to identify putative targets of TSAd activity within alloimmune T regulatory cells. Our hypothesis is that TSAd activity is critical for the generation of alloimmune T regulatory cell function, and further, that the biological effect(s) of TSAd in immunoregulation are mediated via its expression within T cells. We will test this hypothesis in two specific aims in which we will: 1), determine the function of TSAd in allograft rejection, and evaluate its role in regulatory alloimmune responses in vivo, and 2), determine if the functional effect of TSAd in alloimmune regulation is dependent on its expression within T cells. We believe that these studies are ideal for the R21 mechanism, as they will initiate the exploration of a new molecule in the field, and they have significant potential to result in high impact findings. Understanding roles and function(s) for this adaptor in T cells is also likely to be of great clinical importance in transplantation, as they may lead to the identification of targets that shift the immune response from one of immunity/inflammation to one of tolerance. PUBLIC HEALTH RELEVANCE: Organ transplantation is a life saving therapy for individuals with end stage organ failure, but all transplants eventually fail due to a process called chronic allograft rejection. Ongoing research is focused on the understanding of basic mechanisms leading to the development of chronic rejection and mechanisms whereby the alloimmune response causes graft injury. In this research proposal, we plan to determine if an adaptor protein, called TSAd functions in alloimmunity and if we can learn how TSAd-dependent signals can be manipulated in order to achieve long term graft survival in the future.